Method of encapsulating cells in a tubular extrudate

ABSTRACT

Methods and systems are disclosed for encapsulating viable cells which produce biologically-active factors. The cells are encapsulated within a semipermeable, polymeric membrane by co-extruding an aqueous cell suspension and a polymeric solution through a common port to form a tubular extrudate having a polymeric outer coating which encapsulates the cell suspension. For example, the cell suspension and the polymeric solution can be extruded through a common extrusion port having at least two concentric bores, such that the cell suspension is extruded through the inner bore and the polymeric solution is extruded through the outer bore. The polymeric solution coagulates to form an outer coating. As the outer coating is formed, the ends of the tubular extrudate can be sealed to form a cell capsule. In one embodiment, the tubular extrudate is sealed at intervals to define separate cell compartments connected by polymeric links.

REFERENCE TO RELATED APPLICATIONS

This application is a continuation of application Ser. No. 07/461,999,filed Jan. 8, 1990, now U.S. Pat. No. 5,158,881, which is acontinuation-in-part of application Ser. No. 07/121,626, filed Nov. 17,1987, now U.S. Pat. No. 4,892,538.

BACKGROUND OF THE INVENTION

The technical field of this invention concerns methods and systems forencapsulating living cells for the production of biologically activefactors.

There is considerable interest at present in the biologically activeproducts of living cells, including, for example, neurotransmitters,hormones, cytokines, nerve growth factors, angiogenesis factors, bloodcoagulation factors, lymphokines, enzymes and other therapeutic agents.There is also substantial interest in developing new methods and systemsfor producing such biological factors as well as in delivering thesefactors to subjects for therapeutic purposes.

For example, Parkinson's disease is characterized by the degeneration ofthe dopaminergic nigrostriatal system. Striatal implantation of polymerrods which release sustained amounts of a neurotransmitter, dopamine,has been reported to alleviate experimental Parkinsonism in rodents,indicating that the release of dopamine alone in the proper targetstructure may be able to correct this functional deficiency.

In contrast to the limited capacity of a polymeric matrix drug releasesystem, encapsulated dopamine-releasing cells have been proposed as ameans to provide a continuous supply of neurotransmitters. Theencapsulation of neurotransmitter-secreting cells by a permselectivemembrane which permits diffusion of the biological factor may not onlyprohibit the escape of mitotically active cells, but also prevent hostrejection in the case of cross-species transplantation.

A number of researchers have proposed the use of microcapsules--tinyspheres which encapsulate a microscopic droplet of a cell solution --forboth therapeutic implantation purposes and large scale production ofbiological products. However, there are a number of shortcomings to themicroencapsulation approach: the microcapsules can be extremelydifficult to handle (and retrieve, after implantation); their volume islimited; and the types of encapsulating materials which can be used areconstrained (by the formation process) to polymers which can dissolve inbiocompatible solvents.

An alternative approach has been macroencapsulation, which typicallyinvolves loading cells into hollow fibers and then closing theextremities at both ends with a polymer glue. In contrast tomicrocapsules, macrocapsules offer the advantage of easy retrievability,an important feature in therapeutic (especially, neural) implants.However, the construction of macrocapsules in the past has often beentedious and labor intensive. Moreover, due to unreliable closure,conventional methods of macroencapsulation have provided inconsistentresults.

There exists a need for better techniques for macroencapsulation ofcells for both therapeutic implantation and industrial productionpurposes. Encapsulation techniques which can be practiced in a anautomated fashion, and which permit the usage of a wider range ofmaterials and/or provide more reliable closure would satisfy a long feltneed in the art.

SUMMARY OF THE INVENTION

Methods and systems are disclosed for encapsulating viable cells whichproduce biologically-active factors. The cells are encapsulated within asemipermeable, polymeric membrane by co-extruding an aqueous cellsuspension and a polymeric solution through a common port to form atubular extrudate having a polymeric outer coating which encapsulatesthe cell suspension.

In one aspect of the invention, methods are disclosed in which the cellsuspension and the polymeric solution are extruded through a commonextrusion port having at least two concentric bores, such that the cellsuspension is extruded through the inner bore and the polymeric solutionis extruded through the outer bore. The polymeric solution coagulates toform an outer coating. As the outer coating is formed, the ends of thetubular extrudate can be sealed to form a cell capsule. In oneillustrated embodiment, the tubular extrudate is sealed at intervals todefine separate cell compartments connected by polymeric links.

Strings of cell capsules formed in this manner have a number ofadvantages over conventional, cell-encapsulating products. Themulti-compartment form ensures that breaks in the tubular membrane canbe contained to individual cell capsules. Moreover, the design isparticularly advantageous in preparing implantable cell cultures fordelivery of biologically-active factors to a subject for therapeuticpurposes. The string of cell capsules can be coiled, twisted orotherwise deposited in various shapes to provide a dense and compactstructure for implantation. Because the cell capsules are connected toeach other, they can also be readily retrieved, if necessary, followingimplantation. The string-like nature of these products is particularlypreferable over individual spherical microcapsules which typically areretrieved by aspiration (often resulting in a high percentage ofunretrievable capsules and, consequently, inflamation in the subject).

Multi-compartment cell capsule strings can be formed from the tubularextrudate of the present invention by sealing the extrudate at intervalsusing various techniques. For example, the extrudate can be sealed bycompressing it at intervals using mechanical or pneumatic force.Alternatively, the pressure under which the cell suspension or thepolymeric solution is extruded can be modified to collapse the tubularextrudate at intervals and define separate cell compartments. In yetanother technique, the flow of the cell suspension can be interrupted orotherwise impeded at intervals to likewise collapse the tubularextrudate and define cell compartments.

The products of the present invention are particularly well-suited foruse and therapeutic implant devices, such as those disclosed inco-pending U.S. patent application Ser. No. 121,626, "In Vivo DeliveryOf Neurotransmitters By Implanted, Encapsulated Cells" by Aebischer etal. filed Nov. 17, 1987, herein incorporated by reference. In U.S.patent application Ser. No. 121,626, techniques are disclosed forimplanting encapsulated neurotransmitter-secreting cells into a targetregion within a subject's brain, such that the encapsulated cells secreta neurotransmitter and thereby permit constitutive delivery of atherapeutic agent to treat a neurological deficiency, such asParkinson's disease. Alternatively, artificial organs capable ofsecreting other biological factors, such as hormones (e.g., insulin,thymic factors and the like) can also be constructed using the tubularextrudates and multi-compartment cell capsule strings of the presentinvention.

The products of the present invention are also well-suited for use inbioreactors and other in vitro culturing systems, for the production ofdrugs and other useful biological materials. In such applications, cellswhich produce such materials, either naturally, by mutation or byrecombinant design, are encapsuled and allowed to synthesize thematerials which are then collected following secretion into acirculating culture medium. Alternatively, the biological materials canbe accumulated within the cell capsules (e.g., by appropriate control ofthe porosity) and then harvested the materials by removing the strandsfrom the culture medium, lyzing the polymeric membranes and recoveringthe synthesized materials in concentrated form.

The polymeric coating is preferably a semipermeable membrane, that is tosay, a porous structure capable of protecting transplanted cells fromautoimmune or viral assault, as well as other detrimental agents in theexternal environment, while allowing essential nutrients, cellular wasteproducts and cell secretions to diffuse therethrough. As used herein,the term "selectively permeable" or "semipermeable" is used to describebiocompatible membranes which allow diffusion therethrough of soluteshaving a molecular weight up to about 150,000 (Mr).

The permeability of the polymeric coating can be varied by controllingthe viscosity of the polymeric solution, such that upon coagulation, thecoating will form with a network of microchannels to provide diffusionpathways. In one embodiment, this can be achieved by employing awater-miscible solvent as a component of the polymeric solution andmaintaining a pressure differential between the aqueous cell suspensionand the polymeric solution during extrusion. As the tubular extrudateforms, water from the aqueous cell suspension infiltrates into thecoagulating polymer to replace the solvent as the solvent is drivenoutward by the pressure difference. Upon coagulation, the water whichhas infiltrated into the polymeric coating provides a network of pores.The optimal pressure and viscosity will, of course, vary with thesolvent and polymer employed but can be readily ascertained for anyparticular polymer/solvent combination by those skilled in the artwithout undue experimentation.

Various polymers can be used to form the membrane coatings of thepresent invention, including polymers derived from solutions which wouldotherwise be incompatible with the propagation of living cells. Becauseof the unique extrusion process disclosed in the present invention,solvents which would otherwise be toxic are quickly driven away from theaqueous cell suspension during the membrane formation process, therebypermitting the use of many new and potentially useful polymericmaterials. For example, polymeric membranes can be formed frompolyacrylates (including acrylic copolymers), polyvinylidienes,polyurethanes, polystyrenes, polyamides, cellulose acetates, cellulosenitrates, polysulfones, polyacrylonitriles, as well as derivatives,copolymers, and mixtures thereof.

The solvent for the polymer solution will depend upon the particularpolymer chosen for the membrane material. Suitable solvents include awide variety of organic solvents, such as alcohols and ketones,generally, as well as dimethylsulfoxide (DMSO), dimethyacetamide (DMA)and dimethylformimide (DMF), in particular. In general, water-miscibleorganic solvents are preferred.

The polymeric solution or "dope" can also include various additives,including surfactants to enhance the formation of porous channels, aswell as antioxidants to sequester oxides that are formed during thecoagulation process. Moreover, when the cell capsules of the presentinvention are designed for implantation, materials, such asanti-inflammatory agents and cell growth factors, can also beincorporated into the polymeric membrane to reduce immune response orstimulate the cell culture, respectively. Alternatively, these materialscan be added to the multi-compartment cell capsule strands afterformation by a post-coating or spraying process. For example, thestrands can be immersed in a solution which contains ananti-inflammatory agent, such as a corticoid, an angiogenic factor, or agrowth factor following extrusion to post-coat the cell capsules.

Various techniques can also be employed to control the smoothness orroughness of the outer surface of the polymeric coating. In someinstances, a very smooth outer coating can be preferable to reduce scartissue attachment and other immunoreactions during implantation. Such asmooth coating can be obtained by simultaneously immersing the tubularextrudate in a quenchent, such as a bath of physiological saline, or byapplying a flowing, quenchent fluid during the extrusion (e.g., from athird, concentric, outermost bore in an extrusion head assembly).Alternatively, in some applications a rough outer surface with largerpores may be desired, for example, in instances where capillary ingrowthduring implantation is desired, and such a rougher outer surface can beobtained by coagulation in air.

Various cell lines can be encapsulated according to the presentinvention. As noted above, the multi-compartment cell culture stringsare particularly useful for the constitutive delivery ofneurotransmitters, such as dopamine, which is secreted by cells of theadrenal medulla, embryonic ventral mesencephalic tissue and neuroblasticcell lines. PC12 cells (an immortalized cell line derived from a ratpheocromocytoma) are particularly preferred in some applications becauseof their ability to secrete large amounts of dopamine over long periodsof time. Other neurotransmitters include gamma aminobutyric acid (GABA),serotonin, acetylcholine, noradrenaline and other compounds necessaryfor normal nerve functions. A number of cell lines are known or can beisolated which secrete these neurotransmitters. Cells can also beemployed which synthesize and secrete agonists, analogs, derivatives orfragments of neurotransmitters which are active, including, for example,cells which secrete bromocriptine, a dopamine agonist, and cells whichsecrete L-dopa, a dopamine precursor.

In other embodiments of the invention, the encapsulated cells can bechosen for their secretion of hormones, cytokines, nerve growth factors,angiogenesis factors, antibodies, blood coagulation factors,lymphokines, enzymes, and other therapeutic agents.

The aqueous cell suspensions can further include various additives toprotect the cells during the extrusion process or to stimulate theirgrowth subsequently. Such additives can include, for example a nutrientmedium or growth factors which are incorporated into the aqueoussuspension, as well as an anchorage substrate material to enhance cellattachment. The anchorage substrate material can be a proteinaceousmaterial, such as collagen, laminin, or polyamino acids. Alternatively,the cell suspension or the polymeric solution (or both) can include afoaming agent or a blowing agent which can distort the inner surface ofthe polymeric coating to increase the anchorage surface area of thetubular interior.

The products of the present invention can take various forms, includingsimple tubular extrudates as well as multi-compartment cell capsulestrings. The shape of the multi-compartment strings can be tubular,resembling sausages, or nearly spherical, resembling strings of pearls.The maximum outer diameter of the strand with typically range from about0.1 to about 1.0 millimeters. The membrane wall thickness will typicallyrange from about 10 to about 100 microns. The length of the strands canbe varied depending upon the particular application.

In another aspect of the invention, systems are disclosed forencapsulating cells to produce the tubular extrudate andmulti-compartment cell capsule products described above. This system caninclude an extrusion head assembly (e.g., a spinneret or the like)having a first inner bore and a second, concentric, outer bore, as wellas a cell suspension supply means for supplying the aqueous cellsuspension to the inner bore of the extrusion head assembly, and apolymeric solution supply means for supplying the polymeric solution tothe outer pore of the extrusion head assembly. As the cell suspensionand polymeric solution are co-extruded, they form a tubular extrudatehaving a polymeric outer coating which encapsulate the cell suspension.

The tubular extrudate can be sealed at intervals by any one of a numberof mechanisms. In one illustrated embodiment, two wheels with occludingelements on their periphery cooperate in rotation to periodically pinchthe tubular extrudate and thereby seal it. This mechanical compressionsystem can be replaced by a variety of other mechanical or pneumaticcompression systems to seal the tubular extrudate at intervals.

Alternatively, the system can include a flow control means for varyingthe pressure differential between the aqueous cell suspension and thepolymeric solution during co-extrusion. For example, each of thecomponents supply means can include an infusion pump which is operatedby a computer or other control element. In the normal operation, theinfusion pumps are controlled to maintain a pressure differentialbetween the aqueous cell suspension and the polymeric solution, suchthat the polymer solvent is driven outward during coagulation. Byperiodically varying the pressure, the tubular extrudate can becollapsed at intervals to define individual cell compartments. This canbe accomplished, for example, by reducing the aqueous solution pressure.In some instances, it may be preferable to terminate the flow of theaqueous solution entirely and create a vacuum to ensure a complete sealbetween compartments.

Various other techniques can likewise be employed to interrupt the flowof the aqueous solution at intervals and thereby cause the tubularextrudate to collapse and form multiple compartments. For example, aretraction mechanism can be incorporated into the extrusion headassembly for moving the inner bore relative to the outer bore, such thatthe flow of the aqueous solution is interrupted to define separate cellcompartments at intervals.

The systems disclosed herein can further include a quenchent bath forcoagulating the polymeric solution following extrusion, and variousmechanisms for drying the tubular extrudate as it emerges from theextrusion head, including blowers, or evacuation chambers. The extrusionhead assembly can incorporate additional bores to provide multiplecoatings or to deliver a quenchent fluid about the tubular extrudate.The system can also include a sedimentation chamber for the cellsuspension, or an equivalent cell packing mechanism, to increase thecell density within the aqueous cell suspension.

The invention will next be described in connection with certainillustrated embodiments; however, it should be clear that variousadditions, subtractions or modifications can be made by those skilled inthe art without departing from the spirit or scope of the invention.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is an overall schematic diagram of a system for encapsulatingviable cells according to the invention;

FIG. 2 is a more detailed schematic diagram of an extrusion headassembly for use in the system of FIG. 1;

FIG. 3 is a schematic diagram of an alternative extrusion head assemblyfor use in the system of FIG. 1;

FIG. 4 is a schematic diagram of a mechanism for periodically sealing atubular extrudate according to the invention to form a multi-compartmentcell culturing vehicle;

FIG. 5 is a graph showing dopamine release versus time for capsulescontaining dopamine secreting cells produced according to the presentinvention with three different solvent systems;

FIG. 6 is graph showing dopamine release by PC12 cells under normal andpotassium-stimulated conditions at various times following encapsulationaccording to the invention; and

FIGS. 7A and 7B are graphs showing the release of catecholamines forPC12 cells and chromaffin cells respectively.

DETAILED DESCRIPTION

In FIG. 1, a system 10 is shown for producing a tubular extrudate 12according to the present invention, including an extrusion head 14having a first (innermost) bore 16, a second outer bore 18 and,optionally, a third (outermost) bore 20. The system 10 further includesa cell suspension supply 22 and an associated pump 24, a polymersolution supply 26 and an associated pump 28 and, optionally, a flushsolution supply 30 with a pump 32. Additionally, the system can also,optionally, include a outer flowing quenchent supply 34 with anassociated pump 36. All of the pump elements can be controlled manuallyor, preferably, by an automated controller (e.g., a microprocessor) 38.The system 10 can also include a quenchent bath 40, which would normallybe disposed directly below the extrusion head 14 during operation.

When the system 10 is employed to shape the tubular extrudate into amulti-compartment cell capsule string, a sealing means can be employed.One such sealing element 42 is shown in FIG. 1, including two motorizedwheels 44A and 44B which have a series of protuberances 46 whichcooperate during rotation to periodically pinch and seal the tubularextrudate as it passes between the wheels 44A and 44B. Alternatively, aretraction means 48 can be employed to periodically retract the innerbore so as to interrupt the flow of the cell suspension. The effect ofthese retractions is to periodically seal the tubular extrudate andagain form multiple compartments. In yet another alternative approach,the controller 38 can vary the pressure applied by pump 24 (and/or pump28) to create periodic interruptions in the flow of the cell suspension.

In FIG. 2, the extrusion head 14 is shown in more detail, including aninner bore 16 for delivery of a cell suspension and an outer bore 18 fordelivery of a polymeric solution. As the cell suspension and thepolymeric solution are extruded through the common extrusion port 19,the polymeric solution coagulates to form an outer coating about thecell suspension.

In FIG. 3, an alternative extrusion head 14A is shown in more detailcomprising an inner bore 16 for the delivery of the cell suspension, asecond bore 18 (surrounding the inner bore) for delivery of thepolymeric solution, and an outer most bore 20 for delivery of a flowingquenchent fluid, such as saline. In this embodiment, a smooth coatingcan be obtained by simultaneously extruding the cell suspension andpolymeric solution through common port 19 while applying a flowingquenchent fluid during the extrusion (e.g., from the outer most bore 20in the extrusion head assembly 14A.)

In FIG. 4, the sealing element 42 of FIG. 1 is shown in more detail.Motorized wheels 44A and 44B are mounted on opposite sides of thetubular extrudate 12, such that upon rotation protuberances 46 on thewheels periodically come in contact with the extrudate 12 to pinch andseal the extrudate 12 as it exits the extrusion head 14. The wheels 44Aand 44B can be mechanically linked and operated by a conventional motorunder the control of a controller, such as shown in FIG. 1. The resultof the periodic sealing of the extrudate 12 is a multi-compartmentmacrocapsule strand 50 having a polymeric membrane 52 surrounding anencapsulated cell solution 54 with individual cells 56 disposed therein.The individual cell capsules are joined to each another by connectivefilaments 58 where the protuberances 46 of the sealing means 42 hasPinched the extrudate 12.

The invention will next be described in connection with certainillustrative, non-limiting examples:

EXAMPLES

An extrusion system similar that illustrated in FIG. 1 was used,consisting of three electronically controlled programmable infusionpumps, a jet spinneret, two motor-controlled, coaxial wheel systems onthe perimeter of which occluding polytetrafluoroethylene tubes weremounted, and an uptake system.

The macrocapsules were formed by injection of a polymeric solution intothe outer tube of the spinneret. A coagulant, typically the encapsulatedcells in their culture medium, was simultaneously injected in thespinneret inner tube. The encapsulating membrane was formed by adry-jet, wet spinning process, i.e., the fast stabilization of thepolymer solution emerging from the spinneret nozzle by the internalquench medium coupled with further stabilization in a quench bath. Theclosure of the fiber was performed by mechanically squeezing the forminghollow fiber with the coaxial wheel system prior to immersion in thequench bath. Near the spinneret head, the solvent concentration wassufficiently high to allow proper fusion of the fiber wall. Followingeach round of encapsulation, pure solvent was flushed automaticallythrough the lumen of the spinneret to avoid clogging of the nozzle.

PC12 cells, an immortalized cell line derived from a rat pheocromocytomawhich secretes large amounts of dopamine, were cultivated oncollagen-coated tissue culture dishes in RPMI 1640 medium supplementedwith 10% heat inactivated horse serum and 5% fetal calf serum.Dissociated bovine adrenal medullary cells, a non-dividing cell typewhich secretes dopamine, were maintained in DMEM medium supplementedwith 5% fetal calf serum. Prior to encapsulation, the cells wereharvested and loaded at a concentration of 1×10⁵ cells/ml in a 3 mlsyringe. A 15 percent vinylchloride-acrylonitrile copolymer solution ineither dimethylsulfoxide (DMSO), dimethylformimide (DMF), ordimethylacetamide (DMAC) was loaded into a 5 ml glass syringe. Bothsolutions were then coextruded through the spinneret, and the capsuleswere collected in a physiologic saline solution. The capsules wererinsed and placed in individual wells containing the appropriate culturemedia.

Basal and potassium-evoked release of catecholamines was quantifiedunder static incubation conditions by ion-pair reverse-phase highperformance liquid chromatography (HPLC) equipped with electrochemicaldetection at 2 and 4 weeks. Morphological analysis, including light,scanning, and transmission electron microscopy, was performed onrepresentative samples for each time period.

All cell-loaded capsules released dopamine into the medium under basalconditions at all time periods. High potassium treatment increaseddopamine release from both PC12 and adrenal medullary cells. Dopamineoutput by PC12 cells, but not adrenal medullary cells, increased withtime. The increase in dopamine release by the PC12 cell-loaded capsulesover time is believed to be related to cell proliferation within thepolymer capsule. No significant difference in dopamine release could beobserved from PC12-loaded capsules extruded with the three differentsolvent systems (DMSO, DMF, DMAC), which suggests that the encapsulationtechnique of the present invention may prevent cell damage inflicted bysolvents (FIG. 5). Due to the higher pressure of the inner bore system,the solvent was quickly driven toward the outside of the polymer capsulewhich prevented extended cell-solvent contact.

Morphological analysis revealed the presence of small clusters of PC12cells randomly dispersed throughout the lumen of the capsule. At theelectron microscope level, well-preserved PC12 cells, with their typicalelectron-dense secretory granules, could be observed. Cell divisionwithin the capsule space was suggested by the presence of numerousmitotic figures. Although initially coextruded as a cell suspension,adrenal chromaffin cells formed packed aggregates one week afterencapsulation.

FIG. 6 shows the results of an in vitro assay in which PC12 cells wereencapsulated according to the present invention and monitored forrelease dopamine at two and four weeks following encapsulation. Dopaminelevels were measured under both normal (controlled) conditions and alsounder a high potassium stimulation, which is known to inducedepolarization of the cells and, consequently, to increase the secretionof dopamine in viable cells. As can be seen from the graph, there waslittle activity at two weeks; however, at four weeks the encapsulatedcells exhibited dopamine secretions not only under normal conditions butalso exhibited a strong response to the potassium stimulation,indicating that the cells were indeed viable in their encapsulatedstate.

FIGS. 7A and 7B show the result of another in vitro assay in which thesecretions of both PC12 cells and chromaffin cells were monitored fourweeks after encapsulation according to the present invention. Again, thecells were stimulated by high potassium concentrations and the mediumwhile the PC12 cells released only dopamine, the chromaffin cellsreleased a variety of catecholamines. The graft shows the levels ofnoradrenaline, epinephrine, and dopamine.

Due to their fluid dynamics, the macrocapsules extruded in accordancewill the present invention should allow the use of a wider range ofpolymer/solvent systems and can constitute a more efficientencapsulation technique. The results show that immortalized anddifferentiated dopamine-secreting cells will survive inmacroencapsulation. The ability of these capsules to spontaneouslyrelease dopamine over time suggests that polymer encapsulation canprovide an alternative to the transplantation of non-encapsulated ormicroencapsulated dopamine-secreting cells in the treatment ofParkinson's disease.

We claim:
 1. A method of encapsulating viable cells comprising(i)co-extruding an aqueous cell suspension and a polymeric solution througha common extrusion port having at least two concentric bores to form atubular extrudate having a polymeric outer coating which encapsulatessaid cell suspension, wherein said cell suspension is extruded though aninner bore and said polymeric solution is extruded though an outer boreand a pressure differential is maintained between said cell suspensionand said polymeric solution during co-extrusion to impede solventdiffusion from said polymeric solution into said cell suspension andsaid polymeric solution and said cell suspension are chosen so thatcoagulation of said polymeric solution occurs as the polymeric solutionand cell suspension are extruded through the extrusion port; and (ii)sealing said tubular extrudate to form at least one isolated tubularcell compartment.
 2. The method of claim 1 wherein the method furtherincludes sealing the tubular extrudate at intervals to define separatecell compartments connected by polymeric links.
 3. The method of claim 2wherein the step of sealing the extrudate further comprises compressingthe extrudate at intervals to define separate cell compartments.
 4. Themethod of claim 2 wherein the step of sealing the extrudate furthercomprises modifying the pressure under which the cell suspension or thepolymeric solution is extruded, thereby collapsing the tubular extrudateat intervals to define separate cell compartments.
 5. The method ofclaim 2 wherein the step of sealing the extrudate further comprisesimpeding the flow of the cell suspension at intervals to collapse thetubular extrudate and define separate cell compartments.
 6. The methodof claim 1 wherein the extrusion port is in contact with air.
 7. Themethod of claim 1 wherein the extrusion port is in contact with aquenchent.
 8. The method of claim 1 wherein said cell suspensioncomprises cells that secrete a biologically-active factor.
 9. The methodof claim 1 wherein said cell suspension comprises cells that secrete aneurotransmitter.
 10. The method of claim 1 wherein said cell suspensionfurther comprises a nutrient medium.
 11. The method of claim 1 whereinsaid cell suspension further comprises an anchorage substrate material.12. The method of claim 1 wherein said polymeric solution furthercomprises a water-miscible solvent component.
 13. The method of claim 1wherein said polymeric solution further comprises a surfactant.
 14. Themethod of claim 1 wherein said polymeric solution further comprises ananti-inflamatory agent.
 15. The method of claim 1 wherein said polymericsolution further comprises an antioxidant.
 16. The method of claim 1wherein the method further comprises controlling the viscosity of saidpolymeric solution, such that upon coagulation said outer polymericcoating will form a semipermeable membrane about said cell suspension.